Saikosaponin-d regulates angiogenesis in idiopathic pulmonary fibrosis through angiopoietin/Tie-2 pathway.

OBJECTIVE: Idiopathic pulmonary fibrosis (IPF) is considered as a chronic interstitial lung disease with underlying mechanism of IPF remaining unclear, while there are no definitive treatment options. In recent years, scientists have gradually paid attention to the influence of angiogenesis on IPF. Because IPF is a progressive with microvascular remodeling disorder, scientists have postulated that angiogenesis may also be one of the initiating and contributing factors of the disease. Bupleurum is a common natural Chinese herbal medicine with antibacterial, anti-inflammatory, anti-tumor and other pharmacological effects. As the most important active monomer of Bupleurum, Saikosaponin-d (SSd) is a new discovery with anti-pulmonary fibrosis (PF) activity. This study attempts to investigate the role of SSd in the interference of PF through regulation of angiogenesis in IPF through Angiopoietin (Angpt) /Tie receptor 2 (Tie2) pathway. METHODS: Randomly, we allocated C57BL/6 mice into four groups (n = 20 in each group). Afterwards, establishment of IPF model was accomplished via intratracheal administration of bleomycin (BLM, 5 mg/kg), while corresponding drug intervention was given accordingly. On 3rd, 7th, 14th and 28th days after modeling, we performed histopathological examination through staining. Meanwhile, immunohistochemistry (IHC) of PF and the expression of related factors were observed, while Ang/Tie2 pathway was assessed by ELISA with the effect of SSd on angiogenesis related proteins in IPF being explored with IHC and Western Blot technique. RESULTS: Our results showed that SSd could reduce inflammation and PF levels in lung tissue of experimental mice, while levels of angiogenesis-related factors, namely Tie-2, Ang-1 and ANGPT2 (Ang-2), fibrosis- associated factors like Alpha-smooth muscle actin (α-SMA), collagen-I and hydroxyproline in SSd and dexamethasone (DXM) mice were significantly reduced at each time point compared to BLM (p < 0.01). Additionally, we discovered substantial decreased expressions of Ang-1, Ang-2, Tie-2, α-SMA and collagen-I at protein level in SSd and DXM mice at each time point compared to BLM (p < 0.05). Besides, insignificant differences were observed between SSd and DXM groups (p > 0.05). CONCLUSION: This study has demonstrated that SSd could down-regulate the expression of ANG-1, Ang-2 and Tie2 in the Ang/Tie2 pathway, and may reduce lung inflammation and PF in BLM-induced mice via inhibition of angiogenesis.

Dietary Methionine Restriction Alleviates Choline-Induced Tri-Methylamine-N-Oxide (TMAO) Elevation by Manipulating Gut Microbiota in Mice.

Dietary methionine restriction (MR) has been shown to decrease plasma trimethylamine-N-oxide (TMAO) levels in high-fat diet mice; however, the specific mechanism used is unknown. We speculated that the underlying mechanism is related with the gut microbiota, and this study aimed to confirm the hypothesis. In this study, we initially carried out an in vitro fermentation experiment and found that MR could reduce the ability of gut microbiota found in the contents of healthy mice and the feces of healthy humans to produce trimethylamine (TMA). Subsequently, mice were fed a normal diet (CON, 0.20% choline + 0.86% methionine), high-choline diet (H-CHO, 1.20% choline + 0.86% methionine), or high-choline + methionine-restricted diet (H-CHO+MR, 1.20% choline + 0.17% methionine) for 3 months. Our results revealed that MR decreased plasma TMA and TMAO levels in H-CHO-diet-fed mice without changing hepatic FMO3 gene expression and enzyme activity, significantly decreased TMA levels and expression of choline TMA-lyase (CutC) and its activator CutD, and decreased CutC activity in the intestine. Moreover, MR significantly decreased the abundance of TMA-producing bacteria, including Escherichia-Shigella (Proteobacteria phylum) and Anaerococcus (Firmicutes phylum), and significantly increased the abundance of short-chain fatty acid (SCFA)-producing bacteria and SCFA levels. Furthermore, both MR and sodium butyrate supplementation significantly inhibited bacterial growth, down-regulated CutC gene expression levels in TMA-producing bacteria, including Escherichia fergusonii ATCC 35469 and Anaerococcus hydrogenalis DSM 7454 and decreased TMA production from bacterial growth under in vitro anaerobic fermentation conditions. In conclusion, dietary MR alleviates choline-induced TMAO elevation by manipulating gut microbiota in mice and may be a promising approach to reducing circulating TMAO levels and TMAO-induced atherosclerosis.

[γ-aminobutyric acid fortified rice alleviated oxidative stress and pancreatic injury in type 2 diabetic mice].

OBJECTIVE: To investigate the effects of gamma aminobutyric acid(GABA) fortified rice diet intervention on oxidative stress and pancreatic injury in type 2 diabetes mellitus(T2 DM) mice. METHODS: Of the 70 male ICR mice, 10 were randomly selected as blank control group and they were always fed with the normal white rice feed. The remaining 60 mice were fed with high-fat white rice for 9 weeks. They were fasted for 12 h and injected intraperitoneally with streptozocin(STZ) at a dose of 50 mg/kg body weigh for two consecutive days. The control group was injected with the corresponding volume of normal saline. Subsequently, 50 T2 DM mice with successful modeling were randomly divided into 5 groups according to blood glucose, 10 in each group: T2 DM model control group, germinated brown rice positive control group(GABA content is 0. 2 g/kg feed), GABA-fortified rice low, medium and high dose group(GABA content was 0. 02, 0. 1 and 0. 2 g/kg feed respectively) and each target diet was fed for 6 weeks. Oral glucose tolerance test was performed one week before the end of the experiment to observe the hypoglycemic effect of different doses of GABA fortified rice. After the end of the experiment, HE staining was used to observe the morphology of pancreas. At the same time, the redox indicators from plasma and pancreas of reactive oxygen species(ROS), malondialdehyde(MDA), total antioxidant capacity(T-AOC), glutathione peroxidase(GSH-Px), superoxide dismutase(SOD) were examined in each group; The mRNA expressions of oxidative stress-related genes including glycogen synthase kinase-3ß(GSK-3ß), nuclear transcription factor 2(Nrf2), heme oxygenase 1(HO-1) and NAD(P)H: quinone oxidoreductase1(NQO1), insulin secretion related genes including pancreatic and duodenal homeobox 1(PDX-1), mus musculus v-maf musculoaponeurotic fibrosarcoma oncogene family, protein A(MafA), glucokinase(GCK), glucose transporter 2(GLUT2) and the apoptosis associated genes including b-cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax) and caspase-3 in pancreas were assayed by real-time fluorescent quantitative PCR. RESULTS: The intervention of GABA fortified rice could alleviate the improvement of the blood glucose level and the lack of insulin secretion in T2 DM mice and relieve plasma and pancreatic oxidative stress. besides, The intervention of GABA fortified rice could up-regulate the expression of insulin secretion-related genes PDX-1, GCK, GLUT2, inhibit the expression of pro-apoptotic gene caspase-3 and promote the expression of anti-apoptosis gene Bcl-2. There was a dose-response relationship between the above result and the 0. 2 g/kg dose group was the most significant, which achieved similar result to germinated brown rice. CONCLUSION: GABA-fortified rice can significantly improve the plasma and pancreatic redox status of STZ-induced T2 DM mice, regulate the expression levels of oxidative stress-related genes and apoptosis-related genes, thereby protect pancreatic tissue morphology, improve pancreatic insulin secretion and thereby alleviate glucose metabolism.